The present invention relates to a particle analyzing method and device capable of measuring particles such as red blood cells, etc. to obtain various sorts of information on relevant particles, and, in particular to a particle analyzing method and device for measuring the size distribution of the particles.
By a prior art particle analyzing method for analyzing extremely small particles it is known that, as indicated in FIG. 1, particles to be measured 102, are floated in an electrolytic solution such as a sodium chloride solution, etc., and are passed through an orifice 108. Electrodes 104 and 106 are disposed, on either of the orifice 108, to obtain information on the number and the size of the particles 102 by measuring variations in the electric impedance between the two electrodes 104 and 106 during passage of the particles 102 through the orifice (Coulter Counter Method). Refer to e.g. U.S. Pat. Nos. 2,656,508 and 4,348,107. Using the Coulter Counter method variations in the electric impedance are proportional to the volume of the particles and are used to produce a histogram of the particles. However, in order to correctly measure the variations in the electric impedance, it be necessary that the orifice sufficiently long in its axial direction. Further when the concentration of particles in an electrolytic solution containing particles is high, the that a plurality of particles will pass through the orifice simultaneously, increases.
That is, if a plurality of particles 102a and 102b exist in the orifice 108 simultaneously, as indicated in FIG. 2, the information obtained from the impedance at that time is as if there is only one particle and the volume thereof is the sum of the two particles 102a and 102b.
This problem of the simultaneous passage of two particles, called "coincidence", with respect to the information on the number of particles can be corrected mathematically, but similar correction of the information on the volume is not possible. This is because it is not possible to distinguish clearly whether the variations in the electric impedance due to the passage of particles are produced by one particle or by a plurality of particles.
FIG. 3 shows an example of a particle volume histogram when particles having a uniform size at a high concentration are measured by the Coulter counter method. In this figure, the full line A.sub.1 indicates a correct particle volume histogram, i.e. that obtained when no coincidence is produced. The broken line A.sub.2 shows a particle volume histogram obtained in the case where coincidence occurs frequently. In the particle volume histogram indicated by the broken line A.sub.2, individual particles having a particular volume, should appear in the neighborhood of the higher peak but, are distributed in the neighborhood of the second lower peak by the fact that a plurality of them pass through the orifice simultaneously. For this reason correct volume histogram information cannot be obtained.
As one method for solving the problem produced by coincidence, it is conceivable to reduce the concentration of particles. However, when the concentration of particles is reduced, the frequency of the passage of particles through the orifice is reduced, and a long time is necessary for measuring the impedance for an optional number of particles required for obtaining a desired counting precision, making the solution unpractical.
On the other hand, another method of correcting this error is known, where light scattering techniques are used for fine particle analysis. Refer to e.g. JP-A-Sho 60-115858. By this method, although the information on the size of particles can be obtained similarly, it is not possible to obtain any precise information on the volume with respect to the Coulter counter method. This is because measured data are influenced strongly by variations in the refractive index of particles themselves, intensity of light absorption, internal structure and shape. However, by this light scattering method (hereinbelow called light detecting method) it is possible to focus light into a spot at the focal point approximately as large as the particles by utilizing laser light as a light source. That is, since it is possible to make the measured domain as small as a square indicated by a dot-dashed line in FIG. 2, the probability is extremely small that more than two particles exist in the extremely small measured domain described above. Consequently this method has an advantage that the probability of measuring coincidence can be lowered with respect to the Coulter counter method.
As described above, the Coulter counter method and the light detecting method have different advantages.